Over 30% of Americans are obese, creating a medical and economic burden approaching $147 billion annually. The development of effective anti-obesity interventions could lead to significant improvements in obesity and related aspects of human health. Three distinct adipose tissue (AT) types have been identified. White (WAT) depots are used for energy storage, while brown (BAT) and beige (BeAT) depots consume energy to produce heat. Therefore, approaches that lead to enhanced prevalence of activation of BeAT/BAT have emerged as promising anti-obesity interventions. WAT and BAT have distinct phenotypes and genomic identities, with BAT typically consisting of a more extensive vascular network and cells that have smaller lipid droplets, dense and more prominent mitochondrial networks. BeAT is phenotypically and functionally similar to BAT, but is found among predominantly WAT depots. Non-invasive imaging methods, such as PET and diffuse optical imaging, have been used to detect BAT/BeAT in whole animals and humans, but lack in resolution and specificity to AT type; traditional assays are invasive. Thus, the overall objective of this project is to develop minimally invasive, and thus, potentially repeatable, high-resolution optical imaging methods to identify BAT, WAT and BeAT, in order to enable efficient development of anti-obesity treatments. The specific objective of this proposal is the identification and validation of high-resolution endogenous fluorescence and scattering-based quantitative biomarkers of different ATs and their functional state. We propose to assess mitochondrial optical redox ratio and organization, lipid droplet biochemical and morphological characteristics, and vascularization as biomarkers of AT type (Aim 1) in apn animal model that exhibits increased BeAT depots. We will employ immunohistochemical and gene and protein expression assays to validate these optical biomarkers, and we will demonstrate that they can be used to monitor at the cellular level the effects of thermogenic treatment designed to induce BAT/BeAT activation (Aim 2). The expected outcome of these studies is a set of novel optical biomarkers of AT type and functional state that can be used as a powerful aid in the development of effective anti-obesity interventions.